Lyme disease continues to represent a major public health problem, and many aspects of Lyme disease pathogenesis and immunology thus warrant further investigation. Decorin-binding protein A (DbpA), a membrane lipoprotein of Borrelia burgdorferi, has been implicated in subserving the parasitic strategy of B. burgdorferi by functioning as a cell matrix-binding adhesin during mammalian tissue invasion. DbpA also has emerged as the most prominent new human Lyme disease vaccine candidate. However, many features of DbpA, such as its temporal expression, membrane topology, role in B. burgdorferi virulence, and overall utility as a protective immunogen, remain poorly understood. The current study addresses these important information gaps. To this end, the Specific Aims of this proposal are: (1) To examine the temporal expression pattern(s) of DbpA by tick-transmitted B. burgdorferi in the mammalian (mouse) host, (2) To examine the membrane topology of DbpA expressed by tick-transmitted B. burgdorferi in the mammalian (mouse) host, (3) To construct a DbpA-deficient mutant of virulent B. burgdorferi by insertional inactivation of dbpA, with emphasis on examining the role of DbpA expression in B. burgdorferi infectivity, virulence, and disease pathogenesis, and (4) To examine further the overall efficacy of DbpA as a vaccinogen in the mouse model of Lyme borreliosis. Regarding the latter, emphasis will be placed on (a) purifying recombinant DbpA (as immunogen) to preserve its native conformation and (b) tick infestation (challenge route for B. burgdorferi) of DbpA-immunized mice to mimic natural B. burgdorferi transmission. The current proposal represents a comprehensive study of the role of DbpA in both the pathogenesis of Lyme disease and its potential as a human Lyme disease vaccine.